Characterization of insulin receptor carbohydrate by comparison of chemical and enzymatic deglycosylation

Abstract

To characterize the carbohydrate moieties of the insulin receptor on IM-9 lymphocytes, the cells were surface iodinated and solubilized, and the insulin receptors were precipitated with anti-receptor antibody. The precipitates were resuspended, subjected to either enzymatic digestion or chemical treatment with trifluoromethanesulfonic acid and the relative mobilities of the alpha and beta subunits before and after treatment were analyzed by polyacrylamide gel electrophoresis and autoradiography. The results indicate that the alpha subunit possesses primarily N-linked carbohydrate which is both complex (Endoglycosidase F sensitive) and polymannose (Endoglycosidase H sensitive). The beta subunit also contains polymannose oligosaccharide units and has, in addition, a substantial amount of carbohydrate which is removed by chemical treatment but is not susceptible to Endoglycosidase F, suggesting the presence of O-linked saccharides. The apparent molecular weights of the core protein of the mature alpha and beta subunits as determined by gel electrophoresis following complete deglycosylation are 98 kDa and 80 kDa, respectively.